Novel genes involved in the control of hematopoiesis

Abstract

The present invention provides three novel HSC genes designated SCM 3, SCM 26, and SCM 113, the coding regions thereof, the gene products, applications of the genes, DNA constructs, vectors and transformed cells each comprising the gene or a fragment thereof. Methods of using the SCM 3, SCM 26 and SCM 113 polynucleotide and polypeptide sequences are also disclosed.

What is claimed is:  
     
         1 . An isolated nucleic acid molecule comprising a polynucleotide sequence encoding a polypeptide that blocks differentiation of mammalian hematopoietic stem cells, wherein said polypeptide comprises an amino acid sequence selected from the group consisting of: 
 a) an amino acid sequence at least 85% identical to SEQ ID NO:2;    b) an amino acid sequence at least 85% identical to SEQ ID NO:4;    c) an amino acid sequence at least 85% identical to residues 240-543 of SEQ ID NO:4; and    d) an amino acid sequence at least 85% identical to SEQ ID NO:6.    
     
     
         2 . The isolated nucleic acid molecule according to  claim 1 , wherein said polypeptide comprises an amino acid sequence at least 90% identical to SEQ ID NO:2.  
     
     
         3 . The isolated nucleic acid molecule according to  claim 2 , wherein said polypeptide comprises an amino acid sequence at least 95% identical to SEQ ID NO:2.  
     
     
         4 . The isolated nucleic acid molecule according to  claim 1 , wherein said polypeptide comprises an amino acid sequence at least 90% identical to SEQ ID NO:4.  
     
     
         5 . The isolated nucleic acid molecule according to  claim 4 , wherein said polypeptide comprises an amino acid sequence at least 95% identical to SEQ ID NO:4.  
     
     
         6 . The isolated nucleic acid molecule according to  claim 1 , wherein said polypeptide comprises an amino acid sequence at least 90% identical to residues 240-543 of SEQ ID NO:4.  
     
     
         7 . The isolated nucleic acid molecule according to  claim 6 , wherein said polypeptide comprises an amino acid sequence at least 95% identical to residues 240-543 of SEQ ID NO:4.  
     
     
         8 . The isolated nucleic acid molecule according to  claim 1 , wherein said polypeptide comprises an amino acid sequence at least 90% identical to SEQ ID NO:6.  
     
     
         9 . The isolated nucleic acid molecule according to  claim 8 , wherein said polypeptide comprises an amino acid sequence at least 95% identical to SEQ ID NO:6.  
     
     
         10 . An isolated polynucleotide sequence consisting of the complement of the nucleic acid molecule of  claim 1 .  
     
     
         11 . An isolated polypeptide that blocks differentiation of mammalian hematopoietic stem cells, wherein said polypeptide comprises an amino acid sequence selected from the group consisting of: 
 a) an amino acid sequence at least 85% identical to SEQ ID NO:2;    b) an amino acid sequence at least 85% identical to either SEQ ID NO:4 or to residues 240-543 of SEQ ID NO:4; and    c) an amino acid sequence at least 85% identical to SEQ ID NO:6.    
     
     
         12 . The isolated polypeptide according to  claim 11 , wherein said polypeptide comprises an amino acid sequence at least 90% identical to SEQ ID NO:2.  
     
     
         13 . The isolated polypeptide according to  claim 12 , wherein said polypeptide comprises an amino acid sequence at least 95% identical to SEQ ID NO:2.  
     
     
         14 . The isolated polypeptide according to  claim 11 , wherein said polypeptide comprises an amino acid sequence at least 90% identical to either SEQ ID NO:4 or to residues 240-543 of SEQ ID NO:4.  
     
     
         15 . The isolated polypeptide according to  claim 14 , wherein said polypeptide comprises an amino acid sequence at least 95% identical to either SEQ ID NO:4 or to residues 240-543 of SEQ ID NO:4.  
     
     
         16 . The isolated polypeptide according to  claim 11 , wherein said polypeptide comprises an amino acid sequence at least 90% identical to SEQ ID NO:6.  
     
     
         17 . The isolated polypeptide according to  claim 16 , wherein said polypeptide comprises an amino acid sequence at least 95% identical to SEQ ID NO:6.  
     
     
         18 . An antibody that binds to the polypeptide of  claim 11 .  
     
     
         19 . A vector comprising the nucleic acid molecule of  claim 1 .  
     
     
         20 . The vector according to  claim 19 , wherein the vector is a retroviral vector.  
     
     
         21 . A host cell comprising the vector of  claim 19 .  
     
     
         22 . A host cell comprising the nucleic acid molecule of  claim 1 .  
     
     
         23 . A method of increasing the effective dose of hematopoietic stem cells in a mammalian subject, comprising: 
 a) obtaining a population of CD34 +  cells which includes a subpopulation hematopoietic stem cells;    b) introducing into the population of CD34 +  cells a polynucleotide sequence encoding the polypeptide according to  claim 11;     c) obtaining a subpopulation of genetically modified stem cells which overexpress said polypeptide; and    d) administering said subpopulation of genetically modified stem cells to a mammalian subject, wherein the effective dose of the hematopoietic stem cells is increased.    
     
     
         24 . The method according to  claim 23 , wherein the polynucleotide is introduced on a vector.  
     
     
         25 . The method according to  claim 23 , further comprising the step of selecting hematopoietic stem cells either prior to or after genetic modification.  
     
     
         26 . The method according to  claim 23 , further comprising the step of culturing the population of CD34 +  cells either prior to or after genetic modification.  
     
     
         27 . The method according to  claim 23 , further comprising introducing a second polynucleotide sequence encoding a therapeutic gene, an antisense gene or a ribozyme into the population of CD34 +  cells.  
     
     
         28 . A method of increasing the effective dose of gene modified cells, comprising: 
 a) obtaining a population of hematopoietic CD34 +  cells which includes a subpopulation hematopoietic stem cells;    b) introducing into the population of CD34 +  cells a polynucleotide sequence encoding the polypeptide according to  claim 11;     c) introducing into the population of CD34 +  cells a second polynucleotide sequence wherein said second polynucleotide sequence encodes a therapeutic gene;    d) obtaining genetically modified cells wherein said cells are capable of expressing the polypeptide according to  claim 11  and the therapeutic gene, wherein the effective dose of the cells capable of expressing the therapeutic gene is increased compared to wild-type cells; and    e) administering the genetically modified cells to a mammalian subject.    
     
     
         29 . A method of producing the polypeptide according to  claim 11 , comprising: 
 a) culturing a host cell comprising a polynucleotide sequence encoding the polypeptide according to  claim 11  under conditions suitable for the expression of the polypeptide; and    b) recovering said polypeptide from the host culture.    
     
     
         30 . A method of blocking the differentiation of mammalian hematopoietic stem cells in vitro, comprising: 
 a) isolating CD34 +  cells from a source of hematopoietic cells;    b) introducing into the CD34 +  cells a vector comprising a polynucleotide sequence encoding the polypeptide according to  claim 11 , whereby a population of said cells are genetically modified by said vector;    c) culturing the modified CD34 +  cells in the presence of at least one cytokine in an amount sufficient to support growth of the modified cells; and    d) selecting cells in which the encoded polypeptide is overexpressed whereby differentiation is blocked.    
     
     
         31 . The method according to  claim 30 , wherein the mammalian hematopoietic cells are human cells.  
     
     
         32 . The method according to  claim 30  further comprising the step of selecting the CD34 +  cells based on the phenotypes selected from the group consisting of Thy-1 + , CD34 + Thy-1 + , CD34 + Thy-1 + Lin − , and CD34 + Thy-1 + CD38 − , either prior to or after introduction of the vector of step b).  
     
     
         33 . A method of blocking the differentiation of mammalian hematopoietic stem cells, comprising: 
 a) introducing into CD34 +  cells a polynucleotide sequence encoding the polypeptide according to  claim 11;     b) genetically modifying a population of said cells with said polynucleotide sequence;    c) allowing expression of said polynucleotide sequence in said cells; and    d) blocking differentiation of said genetically modified cells.    
     
     
         34 . A method of identifying hematopoietic stem cells or progeny thereof, comprising: 
 a) preparing the antibody of  claim 18;     b) purifying the antibody;    c) exposing a population of the hematopoietic cells to the antibody;    d) allowing said cells to bind with the antibody; and    e) selecting said bound cells.